Solid Lipid Nanoparticles: A Promising Drug Delivery System and their Potential for Peptide and Protein Therapeutics

Abstract

The discovery of peptide and protein therapeutics such as insulin and adrenocorticotrophic hormone in the twentieth century was a breakthrough in drug discovery. However, peptide and protein therapeutics due to their characteristics are predisposed to denaturation and degradation and their delivery and formulation have been a persistent challenge for the biotech and pharmaceutical industry. Their bioavailability is very low mainly due to low gastrointestinal solubility and permeability resulting from low membrane penetration, high molecular weight, proteolytic chemical and enzymatic degradation which altogether urge a compatible drug delivery system. Numerous drug delivery systems with modifiable properties have been synthesized. Solid Lipid Nanoparticles (SLNs) protect the encapsulated peptide and protein therapeutics against first-pass effect and proteolytic degradation, thus enhance drug stability, dissolution rate, absorption and bioavailability. The physicochemical properties of SLNs such as small size, high surface area and surface modification improve their mucosal adhesion, tissue-targeted distribution, controlled drug release and half-life. Besides, SLNs can be encapsulated by both hydrophilic and lipophilic drugs which also offer simplicity of preparation, large-scale manufacturing, biodegradability, biocompatibility, low toxicity, low adverse effects and various drug release profile.

1. Introduction

With the breakthrough of novel techniques in pharmaceutical biotechnologies, molecular pharmacology, and medicinal chemistry in recent decades, new approaches have spawned in both drug discovery and drug delivery – as two sides of a coin – to improve the therapeutic efficiency of some of the already-existing drugs and/or introduce new molecular entities (NMEs) into the market. The past decades have witnessed a booming market for protein and peptide drugs (PPDs), owing to their superior efficiency and biocompatibility compared to their chemical counterparts, “small molecular drugs”.

With the introduction of more advanced biomedical analytical methods, and novel genetic and molecular engineering methods in recent decades peptides and proteins, among all endogenous biomolecules of the body, have been the focus of studies and research sector leading to the recognition of various peptides and proteins as NMEs, large-scale protein production and a better-defined role of peptides and proteins as regulatory components of numerous diseases [1], which eventually rendered them as Active Pharmaceutical Ingredients (API) of a new category and generation of medicinal products – e.g. biopharmaceuticals – for the treatment of diseases [2] and recently as a vaccine [3] turning them into the fastest growing sector in pharmaceutical and biotechnology industry [4].

Peptides and proteins have various physiological functions such as hormones, enzyme substrates, enzyme inhibitors, antibiotics, biological regulators, structural components, cellular signaling factors, growth factors, ion channel ligands, neurotransmitters, and catalyzers, and any disorder/dysfunction in their structure or function leads to serious diseases and pathological conditions such as diabetes [5], dwarfism [6], cystic fibrosis [7], thalassemia [8], or impaired blood clotting [9], among many others [10, 11].

PPDs, due to their intrinsic biological characteristics – poor in-vivo stability, poor membrane penetration, low tissue distribution, and low bioavailability [12, 13]– lack the necessary physicochemical requirements and fail to penetrate through biological barriers, implying the importance of proper nanoparticle systems to maintain their biological integrity with the maximum therapeutic concentration and minimum adverse effect [14].

This chapter discusses PPDs, their historical development, pharmaceutical characteristics, studies for their therapeutic application, formulation challenges, and the importance of systematic preformulation studies for their translation into the clinical setting by solid lipid nanoparticles (SLNs) as a promising system for their delivery. There are numerous studies in the literature on the various methodologies of the synthesis of SLNs [15, 16].

2. The journey of “drug discovery” toward peptide/protein therapeutics

In recent decades, the old paradigm of drug discovery – identification of endogenous active compounds/natural products, followed by chemical modifications to optimize their characteristics in in-vivo models – has switched to the modern paradigm, which relies mainly on target-based drug discovery (Figure 1), i.e. in-vitro screening to identify compounds – also known as “leads” – that bind to and inhibit/activate a biological target and then optimize their pharmacological properties, such as target selectivity, pharmacokinetics, and safety, by changing the chemical structure [17].

Figure 1.

Passive (non-targeted) versus active (targeted) drug delivery; cancer cells overexpressing a specific receptor can be identified and targeted with surface-modified nanoparticles.

Hence, receptors as “drug targets” have been the focus of pipeline studies and research for introducing novel drug candidates [18]. The vast majority of drugs exert their effects by interacting with their receptors, which are macromolecules yielding a biological response upon interaction with a drug molecule, followed by a chain of physicochemical events leading to a particular pharmacological response. However, some drugs act extracellularly without involving a drug–receptor interaction at non-cellular constituents of the body (e.g. neutralization of gastric acid by antacid drugs) by acting through macromolecular components, while the biological effects result from non-specific effects of the chemical properties of the drugs (e.g. alcohol acting by destroying the integrity of the cell through disrupting the cellular constituents) [15].

There are basically four classes of receptor molecules – lipoproteins (or glycoproteins), proteins, nucleic acids, and lipids – where proteins and recently peptides have been the main protagonists for drug discovery [19].

The classical strategy in drug discovery revolved around four concepts: improvement of existing drugs, systematic screening, exploitation of biological information, and rational approaches. However, with the advent of genomics in the 1990s, the classical strategies were doubted and questioned, mainly owing to their limited research and development (R&D) productivity in the pharmaceutical sector to introduce potential drug candidates into the market [20]. With the great progress in the development of new tools to identify targets (e.g. RNA interference), optimize their structure (e.g. X-ray crystallography and computational modeling and screening), and compounds that interact with these targets novel preclinical strategies to identify potential drug candidates were brought up namely as target-based screening, phenotypic screening, modification of natural substances and biologic-based approaches, where the first and second accounted mostly for the innovative medicines and their respective molecular mechanism of action with focusing on target-centric approach and limited use of phenotypic screening in studying receptor-drug interactions in diseases’ pathogenesis.

3. Peptides and proteins as an emerged category of drugs

The isolation of insulin as the first peptide in 1921, brought up the potential of peptides and proteins as therapeutic agents. The first application of peptides and proteins as therapeutic drugs in a clinical setting dates back to the 1920s in the endocrinology and hematology fields with insulin and factor VIII, respectively [21]. Over the upcoming decades, with the technologies used for protein purification and synthesis, structure elucidation, and sequencing, enormous studies have been made on the other natural human hormones such as insulin, oxytocin, vasopressin, and gonadotropin-releasing hormone (GnRH) [12] to explore their efficacy as drug candidates, which led to the approval of the first-ever peptide drug product in the early 1980s and the approval of more than 80 peptide drugs and a total of 33 non-insulin peptide drugs onward [22].

Since then, with the exploitation of novel methods in pharmaceutical biotechnologies [23], peptide and protein synthesis and delivery have become a major source of R&D in the pipeline of pharmaceutical and biotechnology enterprises, with over 240 approved PPDs by the FDA and a variety of potential drug candidates in clinical trials [24, 25]. According to 2018 and 2019 PhRMA reports [26] there were respectively 4751 and 5422 novel biotechnological medicines of various therapeutic categories such as insulin, human growth hormone, monoclonal antibodies, interferons, erythropoietin, biopharmaceuticals, biologics, vaccines, therapeutic blood products (such as IVIG), gene therapy and cell therapy (for instance, stem cell therapies) for more than 100 diseases in human clinical trials or under review by the Food and Drug Administration (FDA) [26] including autoimmune disorders, blood disorders, cancer, cardiovascular disease, diabetes-related conditions, GI disorders, ocular conditions, genetic disorders, infectious diseases, musculoskeletal disorders, neurologic disorders, respiratory disorders, skins diseases, transplantation, antiparasitic diseases, AIDS/HIV, etc. [27, 28, 29, 30, 31].

Most of the studies and research conducted on peptides and proteins have been mostly focused on insulin and different delivery routes for insulin. Nevertheless, the high demands urged the pharmaceutical market to shift from human insulin to animal-derived bovine and porcine insulin products until the introduction of recombinant insulin [32, 33], which paved the way for other synthetic peptides’ synthesis, namely synthetic oxytocin [34], synthetic vasopressin [35], and recombinant human insulin [36, 37] alongside natural peptides.

Peptides and proteins have also been used as targeting moieties and surface ligands (e.g. antibodies) on DDSs to promote drug uptake by cancer cells via receptor-mediated endocytosis [38] and the passage through physiological barriers like the blood-brain barrier (BBB) [15]. This strategy has been successfully used in cancer studies for the design and formulation of chemotherapeutic-loaded DDSs (Figure 2) [39].

Figure 2.

Schematic depiction of cancer cell targeting by nanoparticles modified with receptor- specific surface targeting moieties/ligands.

The aforementioned technologies have eliminated the classical methods of peptide and protein extraction and purification from human and animal cells and tissues and resulted in the production of PPDs in commercial quantities. Besides, they can easily be synthesized with chemical synthesis methods (e.g. Merrifield’s solid-phase peptide synthesis method), in which the amino acid sequence of the peptide of interest can be precisely synthesized at the molecular level [40].

The Nordic Council on Medicines in 1976 presented the ATC system as a “drug substances” classification system, which was adopted and recommended by the World Health Organization (WHO) in 1981 as a classification for all global drug utilization studies. However, contemporary emerging trends along with the renewed R&D strategies prompted a new category of biotechnology-driven drugs with potential therapeutic efficiency and lower costs of formulation and marketing. Biotechnology-driven products use genetically modified living organisms to produce protein or peptidyl products, while other pharmaceutical drugs and small molecule drugs (SMDs) (also known as micromolecules) have a low molecular weight of ≤1000 daltons and size of 1 nm that usually derive from chemical synthesis and could regulate a biological process as a drug, which comprises the majority of medicinal products in the market.

4. Advantages and drawbacks of peptide and protein therapeutics

PPDs, in contrast to the SMDs, offer higher potency, selectivity, and specificity for their extracellular targets (as more than 90% of the PPDs have extracellular targets) [25], have biodegradability of non- to low-toxicity metabolites, therapeutic efficiency at low doses, low drug–drug interactions, lower immunogenicity, and allergic reactions, which in turn facilitates the regulatory procedures for their market approval [41]. The large size and flexible backbone of peptide drugs also render them potent inhibitors of PPIs [42]. Other benefits of peptides and proteins as drugs include biocompatibility, cost-benefit, modifiable in-vivo bioactivity, specific targeting, chemical diversity, higher in-vivo activity, stability, and lower expenses favoring them over other categories of drugs for regulatory approval (higher than 20%) which is twice the rate of small molecules [43].

PPDs overcome issues of source availability and safety (transmission of blood-borne pathogens or prion diseases), function as an alternative to direct extraction methods from inappropriate or hazardous sources (e.g. human urine, vipers), and, with their engineering methods, offer clinical advantages over the equivalent natural products on the market (e.g. faster or slower-acting insulins, modified tissue plasminogen activator (tPA), humanized monoclonal antibodies (Mabs), fusion proteins) [44].

Nevertheless, PPDs due to their larger and more complex molecular structure require more advanced analytical methods (e.g. mass spectrometry) to study their physicochemical properties, molecular structure, and biological activity.

PPDs are highly dependent on the production process to maintain their molecular conformation and biological activity (e.g. non-covalent and covalent forces) of peptides and proteins – as Active Pharmaceutical Ingredients (API) – hence, they are sensitive to environmental factors such as temperature, oxidation, light, non-aqueous solvents, metal ions, ionic strength, high pressure, detergents, adsorption, agitation, and shearing forces, which are all inevitably part of the manufacturing, sterilization, and lyophilization processes and eventually might damage the developing peptide/protein leading to biological inactivation, aggregation, immunogenicity and precipitation [45, 46]. PPDs are also vulnerable to numerous physicochemical properties including hydrolysis, oxidation, racemization, β-elimination, disulfide exchange as chemical instability and denaturation, adsorption to surfaces, non-covalent self-aggregation, and precipitation as physical instability. Stringent conditions should also be maintained for their proper storage to avoid degradation.

Antigenicity and immunogenicity of biotech drugs emerging from the application of specific solvents have been voiced as an issue and challenge that remains to be addressed [47].

5. The demand for peptide and protein drug delivery systems

Only recently peptides and proteins have been considered as therapeutic agents while they had never been considered as potential therapeutic agents [43] mostly due to their protease degradation, metabolic instability, short half-life, challenges of the delivery route, low penetration through biological barriers (e.g. blood-brain barrier [15], GI tract [16]), manufacturing complications and high expenses, which in long-term administration render them unfavorable in terms of processing costs and patient compliance especially with regard to parenteral administration as the majority of peptides (10%) have a very low oral bioavailability.

As “drug discovery” of peptide and protein therapeutics has been increasingly addressing an ever-growing range of medical conditions, the pharmaceutical industry is today more in demand of “drug delivery” technologies for their efficient penetration and distribution across biological barriers. Undesired physicochemical features of PPD remain a serious challenge for formulation scientists in the pharmaceutical and biotechnology sectors and enterprises.

Numerous criteria are involved in introducing parenteral and non-parenteral drug delivery systems for PPDs, namely as bioavailability, therapeutic dose and respective release profile (e.g. controlled, sustained release), clinical demand of the market, general or local delivery, disease pathogenesis, the desired delivery route, duration of treatment, patient convenience and compliance, systemic toxicity, synthesis conditions, and process costs. The classical drug delivery of PPDs was based on the parenteral route of liquid formulations which was the invasive and undesired route in terms of patient compliance eventually leading to further investigations on novel DDSs for non-parenteral and non-invasive delivery routes such as oral, nasal, and pulmonary routes for systemic administration and dermal and ocular for topical administrations.

Hence, in recent decades a variety of nanoparticle systems have been introduced and formulated that can provide the desired release profile, improve bioavailability and biodistribution, and when surface-modified are able to passively or actively deliver the therapeutic agent (Figure 1).

The oral delivery with the most consistent formulation and delivery challenges has been the most investigated route and to address low oral bioavailability novel strategies have been introduced in the course of years namely as chemical modification (lipidization, cationization, PEGylation, prodrug formation, peptide cyclization, and unnatural amino acids substitution), addition of effective agents (absorption enhancers, modulation of pH, proteolytic enzyme inhibitor, mucolytic agents, cell-penetrating peptides), medical devices (biodegradable microneedle-based delivery system, ingestible self-orienting system, intestinal mucoadhesive patches), formulation technology with combinational strategies (Transient Permeation Enhancer® (TPE®), Gastrointestinal Permeation Enhancement Technology (GIPET®), peptelligence technology, ThioMatrix™ technology, transferrin-based recombinant fusion protein technology, Oral sCT (Ostora™) technology, Oramed and Orasome technology, Q-Sphera™ technology, Nano Inclusion technology, Oleotec™, and Soctec™ gastro-retentive technology) [48].

6. Solid lipid nanoparticles

Solid lipid nanoparticles (SLNs) belong to lipid-based drug delivery systems and, since the 1990s, have been under investigation as a substitute to address some of the issues of the other DDSs (Figure 3) [49]. SLNs offer biocompatibility, higher penetration capacity, lipophilicity with no need for surface modification, lower toxicity, simple fabrication, stability, low costs, and industrial-scale production. However, they have a low drug-loading capacity justifying partly the reason behind the small number of SLN-marketed products (Table 1) [23].

Figure 3.

Solid lipid nanoparticles (SLNs) and their constituents.

Advantages

Encapsulation capacity for both hydrophilic and lipophilic drugs; Potential encapsulation with a wide range of therapeutic molecules, such as oligonucleotides, peptides, genes, and superparamagnetic iron oxide particles; Protection of the loaded therapeutic molecule from RES clearance; Poor water solubility favoring the encapsulated substance for controlled and sustained release; Long-term stability and lower toxicity making them applicable for long-term administration; Biocompatibility, easily sterilized, and no need for organic solvents use which might influence the toxicity of the final product; Large-scale industrial production capacity; Modifiable targeting features for tissue-targeted drug delivery

Disadvantages

Encapsulated therapeutic particles export, Gelation predisposition, Low encapsulation efficiency

Table 1.

Advantages and disadvantages of SLNs.

SLNs are formulated only with solid lipids which although limit their encapsulation capacity but gives them more controlled drug release due to limited drug mobility and have been formulated as oral pellets and retard capsules (e.g. Mucosolvan®), as microparticles by spray drying and oral nano pellets. There have been plenty of studies regarding their characteristics and properties [50].

Based on the desired release profile, there are three types of SLNs where the therapeutic molecules can be incorporated in the core, matrix, or attached on the surface (in the case of a high surface/volume ratio) [51]. The latter results in a longer half-life, systemic circulation, and increased mean residence time (Figure 4, Table 2) [52].

Figure 4.

Different modes of drug encapsulation in solid lipid nanoparticles.

Model	Drug loading site	Drug release pattern
Homogenous matrix of solid solution	Homogeneous drug dispersion in the lipid matrix of the particles	Diffusion from the solid lipid matrix and/or by degradation of lipid matrix in the GI tract
Drug-enriched shell	Drug concentration on the outer shell of the nanoparticles	Burst release is modified by varying the formulation conditions: production temperature (preferably cold homogenization) and surfactant concentration
Drug-enriched core	Drug concentration in the core of the nanoparticles	Prolonged drug release

Table 2.

Models of drug incorporation for the lipid nanoparticles.

SLNs are composed of different lipids and surfactants/co-surfactants for solidness at various temperatures and low melting points. The choice of lipids, surfactants, and the composition of SLNs (the solid core: 0.1–30% w/w, surfactants: 0.5–5% w/v) determines release profile, drug encapsulation, stability over time, surface charge, polydispersity index, size, and physicochemical features.

Crystallization process during synthesis leads to low encapsulation efficiency of SLNs limiting the internal space of the lipid core for therapeutic substance encapsulation [53]. The highly ordered crystalline structure of the lipids in an SLN has been recently studied with a detailed description of the internal and external structure of SLN [51].

Since their introduction in the 1990s, SLNs have been used as DDSs in various therapeutic fields including anticancer therapies, antimicrobials, central nervous system (CNS) diseases and/or disorders, site-specific treatments, and various conditions/diseases.

Numerous drugs with large and small lipophilic molecules, high polarity, hydrophilic, and hydrophobic characteristics have been efficiently encapsulated into SLNs proving the versatility of these nanoparticles. SLNs can also be surface-modified with peptide/protein-based targeting moieties and surface ligands (e.g. antibodies) for active targeted drug delivery toward receptor-overexpressing cancer cells (Figure 2) [38].

7. Biological barriers and peptide and protein drug delivery by SLNs

7.1 Parenteral route

Although an invasive route, parenteral drug delivery has been the most common route of PPD administration as it avoids the challenges of other routes, especially the oral route [54]. The intravenous (I.V.) route basically uses two separate bodily systems – the blood system and lymphatic system – for the drug’s general distribution and in-site deposition with defined pharmacodynamics properties. The uptake of nanoparticles by the reticuloendothelial system (RES) and their transfer to the spleen and kidney have been a challenge, hence the application of polyethylene (PEGs) and “stealth technology” to prolong nanoparticles blood circulation time [55, 56, 57]. Peptide and protein drug delivery through the lymphatic system has also been voiced as an undesired route by some studies as the delayed time of distribution might result in peptide and protein enzymatic degradation [58, 59]. The transcellular pathway of nanoparticles leads to their uptake by M cells and transport to the lymph [60].

Furthermore, extensive preformulation studies are required for in-vivo assays of PPD parenteral delivery as they have larger molecular sizes than their chemical drug counterparts which is critical in the design of an effective delivery method for their vascular penetration. The anatomical structure of endothelium (i.e. continuous, fenestrated, and sinusoidal) varies in different organs hence rendering them permeable only to peptides and proteins of a specific molecular size [61]; particles >7 μm in size accumulate in the lung capillaries, while particles of 0.1-7 μm clear through the RES (lowering half-life of peptides); while particles <0.1 μm are collected in the bone marrow [62]. Nevertheless, the large molecular size of PPDs imparts them a good steric and electronic complementarity between ligands, which is the most important prerequisite for binding.

The other drawbacks of the I.V. route include unfavorable patient compliance, variable clearance (few minutes to few days), high risks of vascular extravasation and undesired general distribution, repeated injections for the required therapeutic efficacy leading to local tissue necrosis/phlebitis [63, 64], which altogether might result in severe adverse effects. Besides, I.V. preparations are required to be sterile in order to avoid septicemia, thromboembolism, and thrombophlebitis [62].

The subcutaneous (S.C.) and intramuscular (I.M.) routes have been also investigated for PPDs with the former for vaccines. Based on studies [64], the S.C. route offers a 100% bioavailability for PPDs resulting from different factors such as molecular weight, site of injection, local injection site activity, and pathological conditions. Following S.C. administration, PPDs are absorbed based on their molecular weight; high molecular weight drugs (<16,000 Da) either through the endothelial cells of vessels to capillaries or the local lymphatic system to the thoracic duct and general blood circulation, while small molecular weight ones through the local capillaries [64].

7.2 Non-parenteral routes

Non-parenteral delivery of peptide and protein drugs has been investigated extensively as a non-invasive route to address the issues associated with the parenteral route, patient compliance above all. Oral delivery has been the focus of the majority of studies while there have also been numerous studies investigating nasal, pulmonary, transdermal, and ophthalmic routes for PPDs [65, 66, 67, 68, 69, 70, 71, 72, 73]. Mucosae which have been neglected for drug delivery seem to be a promising approach for drug absorption, especially efficient for biomolecules of large size and molecular weight [65, 74]. The advantages of mucosal surfaces for drug delivery over skin and GI tract can be named as fewer biological barriers to pass for systemic diffusion, rapid absorption, and evading hepatic first-pass effect. However, one practical challenge of mucosae is related to the preparations that are formulated for long-term and local treatment.

Oral drug delivery, despite being a desired route in terms of patient compliance and convenience, faces numerous challenges in formulating DDSs to maintain the biological and chemical integrity of PPDs while distributing through barriers.

The first and foremost challenge of oral delivery is the low oral bioavailability (1%) [75] and subsequent short half-life (<1%) resulting mainly from enzymatic degradation and poor intestinal penetration of PPDs which is claimed to have been increased to 30–50% by pharmaceutical enterprises [76, 77]. Enzymatic degradation results from the susceptibility of PPDs to stomach’s salt-laden pepsin, pancreatic proteases (trypsin, chymotrypsin, elastase, and carboxypeptidase A and B), intestinal brush-border peptidases (endo-, amino- and carboxypeptidases) [65] and intracellular enzymes (cytosolic and lysosomal peptidases) [78] which eventually lead to physical instability [79], aggregation, adsorption, denaturation, enzymatic degradation, poor intestinal penetration, short plasma half-life, and immunogenicity [80, 81]. Poor intestinal penetration and the subsequent low blood absorption and diffusion result from large molecular size, hydrophilicity, charge, and relatively high molecular weight (>500 Da) of PPDs.

PPDs due to their larger molecular size require specific epithelial transporters for their general blood distribution [82]. Their large molecular size allows for specific drug-target interaction with binding pockets that are not normally available to small molecular drugs. These targets are part of intracellular protein-protein interaction networks, which have been recognized in numerous diseases. PPDs in order to interact with such targets must penetrate cells; however, most of them are known to have extracellular targets [83] and are parenterally administered; hence, cellular penetration is not their ordinary route as it is for mucosal surfaces. Currently, the main obstacle to the oral administration of these novel categories of drugs for their maximum therapeutic effects could be addressed as the penetration through intestinal and target cellular membranes. The GI tract is covered with different types of cells (enterocytes, M cells, goblet cells, and paneth cells) and their respective target cells, which allow for broad surface modification of DDSs for passive or active targeting.

The mucous layer covering the epithelium of the GI tract is another barrier that is 400–450 μm thick in some parts of the GI tract and 100–200 μm in the duodenum and jejunum. Mucus is composed of different bodily secretions, including mucins and glycoproteins of high molecular weight, which, owing to the presence of the sialic and sulphonic acid functional groups, possesses negative charges, enabling it to interact with positively-charged peptides and functional groups, which in turn lowers their diffusion rate and absorption.

Moreover, the epithelial cell monolayer membrane of the GI tract, tight junctions, and efflux proteins such as P-glycoprotein aggravate the condition of low oral bioavailability and permeability [84, 85]. PPDs surviving physical and GI enzymatic degradation when absorbed in the general circulation are still subject to liver’s first-pass effect enzymatic metabolism, enzymatic degradation by plasma, and clearance via the kidneys.

Nevertheless, the oral route is non-invasive, painless, easy to self-administrate, has minimum risk of cross-contamination, high patient convenience/compliance, outpatient feasibility, and cost-benefit (no need for sterile manufacturing) [86]. Besides, the oral route does not face the drawbacks of I.V. route: drug extravasation from blood, catheter-related infectious complications, and thrombosis, and is expensive and invasive, especially for chronic conditions.

In recent years, the nasal route has been studied extensively for PPD delivery as an alternative to other routes as it evades the first-pass effect and GI enzymatic degradation, especially as a “shortcut” for CNS diseases/disorders by by-passing BBB and distributing through the olfactory nerve (cranial nerve І) and trigeminal nerve (cranial nerve V) [15, 87]. The nasal route has been promising owing to porous epithelial layers, large surface area, microvilli, and highly vascularized mucose, which collectively favor quick general circulation and distribution of drugs [88, 89].

The governing factors for PPD penetration in intranasal tissue are molecular weight, lipophilicity, and dissociation rate, hence the low penetration of PPDs [90]. The nasal mucosa, due to its vast lymphoid tissue, is considered a desirable site for vaccine delivery than other delivery routes as both local and systemic immune systems can be stimulated [90], especially against respiratory infections, with the nasal mucosa as the first site of contact with the causative pathogens/antigens. One study on respiratory syncytial virus (RSV) proved higher immunization with nasal vaccine delivery than intramuscular or oral vaccination [91].

Pulmonary drug delivery has been increasingly under investigation in recent years as it can be exploited to address both local (e.g. asthma, chronic obstructive pulmonary disease (COPD), cystic fibrosis, pulmonary arterial hypertension, tuberculosis, lung cancer, etc.), and systemic diseases/conditions (e.g. cancer, diabetes, acute pain, immune deficiencies, autoimmune diseases, infections, etc.) [92].

The major challenges of the pulmonary route are first the highly-evolved defensive system of the respiratory tract with its mucociliary clearance for exhaling exogenous materials or depositing and deactivating them [93], and second the use of inhaler devices such as pressurized metered dose inhalers (pMDIs) and dry powder inhaler (DPIs) for the efficient therapeutic regimen [94], which partly justify the slow progress of studies and innovations of the pharmaceutical and biotech industry in this sector.

Lungs with alveolar epithelium, vast vasculature system, and large surface area are desirable sites as they allow for high and rapid absorption of poorly water-soluble drugs with low oral bioavailability [95, 96, 97]. Since the extra- and intra-cellular enzyme activity of lung cells is low, drug degradation is low which provides a higher extension of absorption even for drugs with low absorption rates [98]. However, based on studies peptide/protein adsorption through the pulmonary route is quick leading to quick peak serum levels (which is undesirable when the slow-release profile is required) [99, 100] rendering the pulmonary route a desirable route only when low doses of PPDs are required causing no adverse effects at high peak serum levels.

Dermal and transdermal delivery have also been studied as an area of great interest for topical dermal treatment and other general conditions [71]. Although the stratum corneum, the outermost layer of skin, is the major barrier responsible for low skin penetration, it also functions as a defensive barrier against exogenous particles such as microorganisms [101] and drug delivery systems. However, due to the high amount of lipid in the stratum corneum, lipid-based nanoparticles have demonstrated great biocompatibility and biodegradability for drug delivery [102], as quickly bind to the surface of the skin and facilitate lipid exchange between the stratum corneum’s outer layers [103, 104]. Lipid-based nanoparticles protect the encapsulated drug from bodily chemical degradation, modulate the release profile of interest, and allow for an occlusive effect by an adhesive lipid film formation [105].

Ocular drug delivery has also been under investigation for topical ophthalmic treatment. However, eyes due to their unique anatomical structure (e.g. blood-retina barrier, corneal epithelium) and physiological mechanisms (e.g. the short drug residence time) pose a serious challenge for the local delivery of antibiotics, plasmids, anti-inflammatory, and immunosuppressive agents [106]. Based on studies, the anionic nature of the ocular mucosae can contribute to the development and adhesion of positively charged DDSs on the cornea by increasing drug residence time [106, 107].

8. Protein and peptide drug delivery

Due to the hydrophilic properties of SLNs dispersed phase technologies have been employed for their synthesis. The encapsulation of proteins in the lipophilic matrix of the solid lipid core results in the partition of the aqueous phase during formulation hence surfactants (e.g. emulsion and stabilizers) are used [108]. In one study [109] lyophilic ion coupling improved and facilitated encapsulation of leuprolide and insulin within SLNs where stoichiometry of the ion pair was employed for the former molecule. Various peptides and proteins with different properties have been used for cancer studies implying the multifunctionality of SLNs for both hydrophilic and hydrophobic drugs [110].

The first investigation on peptide drug encapsulation in SLNs was done to incorporate LHRH and thymopentin [111], followed by other studies on insulin with an encapsulation efficiency of about 80% [112]. In another study [113], positively and negatively charged lipid nanoparticles were encapsulated by protein antigens (HBsAg) for intranasal immunization against hepatitis B. So far, different strategies have been employed to increase the bioavailability of peptide and protein therapeutics including:

• Co-administration of enzyme inhibitors such as aprotinin (natural inhibitor of trypsin) [113], ethylenediaminetetraacetic acid (EDTA) [114], sodium glycocholate, camostat mesylate [113, 115], or bacitracin [116]);

• Absorption enhancers such as low molecular weight surfactants, bile salts, calcium ion chelators, or cyclodextrins [117, 118];

• Altering the gastrointestinal retention time using mucoadhesive polymers such as chitosans [119];

• And peptide/protein conjugation to a suitable nanosystem [120].

Among numerous DDSs, SLNs seem a very promising system and their properties mentioned in this chapter offer the four abovementioned opportunities for drug delivery of peptide and protein drugs.

9. Future perspectives and conclusion

After the introduction of insulin as the first peptide drug in the 1920s the progress of marketing peptide and protein drugs has been very limited hence numerous approaches were introduced. The information presented in this chapter provided concepts regarding SLNs and their promising characteristics as a DDS for peptide and protein drug delivery. However, further studies are still required for their translation into clinical settings as there are currently no SLNs for peptide and protein drug delivery in clinical evaluation stages and trials, hence more time is expected for their marketing in the pharmaceutical sector. Proof that they are still in their initial stages of research can be seen in the literature, of which most are in-vitro and in-vivo and limited to research. The majority of SLN reports are based on experimental drugs and are not transferable to clinical trials. However, the studies have demonstrated positive results for their application as they are biocompatible and can be encapsulated by a variety of drugs and also can improve the efficacy and pharmacokinetic profile of the encapsulated drugs. Their limiting factors for their marketing still remain to be addressed namely large-scale manufacturing processes, sterilization, tailoring strategies, and stability issues. The hydrophobic constituents of SLNs render them a favorable scaffold for the encapsulation of hydrophobic and lipophilic drugs where the pharmaceutical market and trends are increasing for the latter.